Estudo cinetico da invertase livre e imobilizada em alginato de calcio

AUTOR(ES)
DATA DE PUBLICAÇÃO

1989

RESUMO

In this work special emphasis was given to the study of the kinetics of sucrose hydrolysis by invertase. The enzyme was used in free as well immobilized in calcium alginate forms. The kinetic parameters ware determined considering the initial rates of reaction, in order to avoid the iinhibition phenomena caused by the products glucose and fructose. The pH and temperature effects on enzymatic activity, as well as the thermal stability of enzyme were studied. The parameters for the free enzyme were determined at 4-0°C and 60°C. Three assumptions related to the "physical effect" on the kinetic of substrate transformations were considered: Two of them were related to the concentration of free water and the other one was related to the molecular diffusivity of sucrose. The intrinsic parameters of the immobilized enzyme were determined at 40°C i n a continuous stirred tank reactor, with standard geometric relations. The effective diffusion coefficients of sucrose, needed for the determination of the kinetic parameters for the immobilized enzyme were determined by two methods: The first one, used the theory of semi infinite plate in contact with a liquid phase with constant concentration of sucrose, at unsteady state and, the second, used gel spheres, initially free of sucrose, that were immersed in a fini te volume of solution of known sucrose concentration, at unsteady state. The optimum conditions of enzymatic reaction were pH 4,0 – 4,5 and 60ºC in both free and immobilized forms. In terms of thermal stability the enzyme was stable at temperature lower than 40 C. But, the immobilized enzyma was less stable than the free form. The activation energies obtained for inversion reaction were found to be 7853 and 10258 cal/mol. While the deactivation energies were 141171 and 116883 cal/mol for the free and immobilized forms, respectively. Better agreement for the kinetic model , for the initial rates of reaction, was obtained when the kinetic model of substrate inhibition was considered, corrected by a factor related to mass diffusivity, Values for Michaelis-Menten (Km) and the inhibition (Ki) constants were respect!vely, 0.0506 mol/l and 0. 309 mol/l at 40° C and 0.0557 mol/l and 0.377 mol/l at 50°C. On the other hand, the classic model of Michaelis-Menten fitted well the results for immobilisted enzyme, either in terms of apparent or effectives constants. The inhibition phenomena of substrate were not observed neither diffusional effects, as verified for free enzyme, up to a substrate concentration of BOO g/1. The intrinsic value of Km for immobilized enzyme was 0.031M and V was found to be 60% of the observed Vmax for the free enzyme, in the same enzymic concentration. The sucrose effective diffussivity in the gel was 0.5 .10-5 cm2 /s, corresponding to 67% of the sucrose diffusity in solution.

ASSUNTO(S)

enzimas imobilizadas

ACESSO AO ARTIGO

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